Bioavailable composition of natural and synthetic HCA

ABSTRACT

The invention relates to a composition comprising hydroxycitric acid (HCA) in combination with either one or both of garcinol and anthocyanin, and its use as a weight-loss therapy in animal subjects, preferably humans. The therapeutic effects for the composition observed in murine and human studies include a reduction in total body weight and body mass index, a reduction in body fat, an increase in lean body mass and content of body water, and a reduction in perceived appetite level. Another composition for use in weight-loss therapy is also described relating to forskolin in combination with either one or both of garcinol and anthocyanin. The anti-oxidant properties of garcinol are described as being enhanced in the presence of HCA and anthocyanin, and the combination of HCA, garcinol and anthocyanin is also shown to exert greater citrate lyase inhibiting properties than either compound alone. Methods of obtaining HCA, garcinol or anthocyanin, or a composition containing all three compounds, are described.

CROSS-REFERENCE TO RELATED APPLICATION

This application is a National Stage entry of International ApplicationNo. PCT/US01/41748, filed on Aug. 17, 2001, which claims priority under35 USC §1.119(e) to provisional Application Ser. No. 60/225,821, filedon Aug. 17, 2000, the entire specification, claims and drawings of whichare incorporated herewith by reference.

BACKGROUND OF THE INVENTION

Hydroxycitric acid is an alpha-hydroxy tribasic acid(1,2-dihydroxypropane-1,2,3-tricarboxylic acid) with two asymmetriccenters, hence the formation of two pairs of diasteroisomers or fourdifferent isomers: (−)hydroxycitric acid (I), (+)hydroxycitric acid(II), (−)allo-hydroxycitric acid (III), and (+)allo-hydroxycitric acid(IV). (1–2) The (−)hydroxycitric acid (HCA) isomer is found in the rindof Garcinia cambogia fruit (fam. Clusiaceae). (1–2) This isomer has beenshown to be a potent linear competitive inhibitor of ATP citrate lyaseenzyme, in vitro, demonstrating a much greater affinity for the purifiedenzyme than its natural substrate citrate as well as the otherstereoisomers of hydroxycitric acid. (1–2) The biological importance ofATP citrate lyase is as a citrate cleavage enzyme which catalyzes theextramitochondrial cleavage of citrate to acetyl CoA and oxaloacetate,and facilitates the biosynthesis of fatty acids. The reversibleinhibition of citrate lyase by (−) HCA may lead to the reduction offatty acids synthesis and lipogenesis. These effects have been measuredand demonstrated in vivo following the oral, intravenous orintraperitoneal administration of (−)hydroxycitrate to experimentalanimals. (3) When (−) HCA was given orally before the feeding period,the animals fed (−) HCA consumed less food and their hepatic synthesisof fatty acids and cholesterol was significantly diminished as comparedto the untreated controls. (3–4) The observed decrease in food intakemay be only one of the factors responsible for the (−) HCA promotedweight loss, because experimentation with rats fed (−) HCA showed weightloss with no decrease in cumulative food intake. (5) It seems that thepotential mechanism of weight loss with (−) HCA may include an energyexpenditure component, the nature of which remains undetermined. (5)This mechanism of energy expenditure, decreased lipogenesis, and thereduction in food intake in (−) HCA-treated animals may result in lossof weight and total body fat content. (6)

Although the potential of (−)HCA as a weight lowering compound has beenrecognized since the 1970's, only few clinical studies have beenconducted with this compound. (7–12). These few studies examiningHCA-mediated prevention of excess body fat, resulted in contradictoryresults, most likely due to HCA being poorly bioavailable in the cytosolof a target cell. In one clinical study of HCA, a controversial highfiber diet was used. The use of a high-fiber diet in combination withHCA may reduce gastrointestinal absorption of HCA, since high-fiberdiets are known to reduce absorption of many nutrients andmicronutrients. This issue becomes critical with HCA because itsreported efficacy in inhibiting the intracellular enzyme, adenosinetriphosphate (ATP)-citrate-lyase, depends entirely on the presence ofHCA inside the target cell.

In their U.S. Patent, the Inventors addressed an important issueregarding the bioavailability of the HCA compounds. The U.S. Pat. No.5,783,603 patent described a manufacturing process leading to a uniquestructure for a potassium salt of HCA, which facilitated its transportacross biological membranes, effectively delivering more HCA into thecytosol for the competitive inhibition of ATP citrate lyase. Althoughthe '603 patent related to an HCA compound having considerably improvedbioavailability, its bioavailability was still relatively inefficient.For example, an in vitro study done on hepatic cells, indicates that 5mM of extracellular potassium HCA could inhibit ATP citrate lyase.However, only 0.5 mM of potassium HCA is actually needed in the cytosolto effectively inhibit ATP citrate lyase. Therefore, a 10-fold excessamount of potassium HCA is needed outside of the target cell in order toachieve a concentration of 1/10 that amount in the cytosol. This findingof relatively poor bioavailability of HCA, was confirmed in pre-clinicalexperiments (14), and points out the need to further improve thebioavailability and efficacy of HCA.

Garcinol, like HCA, is isolated from Garcinia sp. fruit rind, and itexhibits anti-oxidant and chemoprotective properties (15). In oneexperiment, rats fed a garcinol diet (0.01% and 0.05%) showed asignificantly reduced development of azoxymethane (AOM)-induced colonicaberrant crypt foci (ACF) as compared to control animals. Feeding ofgarcinol significantly elevated liver glutathione S-transferase, quinonereductase activities, suppressed O2- and NO generation and expression ofiNOS and COX-2 proteins. These findings suggest a possiblechemopreventive mechanism of garcinol.

Garcinol and isogarcinol were evaluated for their antibacterial activityagainst methicillin-resistant Staphylococcus aureus (16). Thesecompounds showed a minimum inhibitory concentration at 3.1–12.5micrograms/ml, or nearly equal to that of the antibiotic, vancomycine.

In 1981, Krishnamurthy et al. (17) reported the isolation of garcinol,and its colorless isomer, isogarcinol, from Garcinia indica. Theirstructures were proposed on the basis of chemical and spectral data.Garcinol, C38H50O6, m.p. 122(o), crystallized out from the hexaneextract of the fruit rind of G. indica as yellow needles (1.5 percent).The UV spectrum of garcinol suggested that the 1,3-diketone system isconjugated to the 3,4-dihydroxybenzoyl moiety. The IR spectrum of thetrimethyl ether showed the presence of a saturated carbonyl group (1727cm−1) and two oe,beta-unsaturated carbonyl groups (1668 and 1642 cm−1).

Isogarcinol was isolated by column chromatography of the extract. Itsidentity was established by mixed m.p. and by comparison of LN, IR, andPMR spectra. The IR spectrum of isogarcinol indicated the presence ofsaturated carbonyl group (1715 cm−1), an aroyl group (1670 cm−1) and anoe,beta-unsaturated carbonyl group (1635 cm−1).

Rao et al. (18) reported the isolation of cambogin (33H55O6), m.p.242–244, from the latex of Garcinia cambogia tree. The structure wasconfirmed by UV, IR and NMR studies. UV: 231–234, 275–278 and 305–313;IR (Kbr): 1720 (saturated carbonyl), 1680 and 1642 cm−1 (unsaturatedcarbonyl and double bond). Besides cambogin they also reported theisolation of camboginol and related its structure to cambogin.

In 1982, N. Krishnamurthy et al. (19) isolated anthocyanin pigments fromthe fresh red ripe fruits of Kokam (Garcinia indica). The rind portionwas separated from the rest of the fruit and was macerated in a blenderusing methanol containing one percent HCl for three times. The extractswere combined, filtered and concentrated in vacuo at 30° C. Paperchromatography of the Kokam pigment extract showed two anthocyaninbands. The slower moving band was designated as B1 and the other B2. Thetotal anthocyanin concentration was estimated to be 2.4 percent on a dryweight basis; the ratio of B1 to B2 is 1:4.

Anthocyanin B1 was identified as cyanidine-three-glucoside by chemicaland spectroscopic studies. This compound on hydrolysis gave cyanidingand glucose. The UV spectral maximum (527 nm) of the glycosides shiftedto 567 nm with aluminium chloride indicating that 3′- and 4′-hydroxylgroups of the cyaniding are free. The structure was confirmed by directcomparison with a sample of cyaniding-three-glucoside obtained frommulberry.

Anthocyanin B2 was identified as cyaniding-three-sambubioside. Thisanthocyanin on complete hydrolysis gave cyaniding, glucose and xylose.The spectral data suggested that B2 is a three-substituted glycoside ofcyaniding. Hydrogen peroxide hydrolysis removed the disaccharide fromthe pigment which on further acid hydrolysis gave glucose and xylose.The structure was confirmed by direct comparison with a sample ofcyaniding-3-sambubioside isolated from Roselle.

The present invention is based on the unexpected finding that combiningHCA with natural compounds obtained from Garcinia sp. plant includinggarcinol (polyisoprenylated benzophenone) and/or anthocyanin compounds,results in not only an enhancement of the biological activity of HCA butalso that of garcinol and/or anthocyanin.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1: Trendlines of the percentage of subjects who experienced weightloss.

FIG. 2: Average percentage of study subjects who experienced weightloss.

FIG. 3: Percentage of weight loss in groups C and NC in consecutivestudy time intervals.

FIG. 4: Percentage of weight loss in group C and NC at consecutive studytime intervals.

FIG. 5: Average weight loss (lbs) in group C and NC at consecutive studyintervals.

FIG. 6: Average percentage body mass index (BMI) change in groups C andNC at two study time intervals.

FIG. 7: Percentage of body mass index (BMI) change in group C and NC attwo study time intervals.

FIG. 8: Percentage body weight change in group C subjects.

FIG. 9: Percentage body weight change in NC group subjects.

FIG. 10: Percentage lean body mass (LBM) change in group NC subjects.

FIG. 11: Percentage lean body mass (LBM) change in group C subjects.

FIG. 12: Percentage fat loss in group NC subjects.

FIG. 13: Percentage water change in group C subjects.

FIG. 14: Percentage water change in group NC subjects.

FIG. 15: Self assessed appetite levels in group C and NC in consecutivetime intervals.

FIG. 16: Self assessed energy levels in groups C and NC in consecutivestudy time intervals.

FIG. 17: Pulse rate in groups C and NC in consecutive time intervals.

FIG. 18: Systolic blood pressure in groups C and NC in consecutive studytime intervals.

FIG. 19: Diastolic blood pressure in groups C and NC in consecutivestudy time intervals.

DETAILED DESCRIPTION OF THE INVENTION

The invention relates to a composition comprising hydroxycitric acid(HCA) in combination with either one or both of garcinol and anthocyaninand having the effect of increasing bioavailability of HCA in thecytosolic compartment of a cell.

Another object of the present invention, involves the use of theinventive composition comprising HCA in combination with either one orboth of garcinol and anthocyanin, as a weight-loss therapy in animalsubjects, preferably humans. The therapeutic effects for the compositionobserved in murine and human studies include a reduction in total bodyweight and body mass index, a reduction in body fat, an increase in leanbody mass and content of body water, and a reduction in perceivedappetite level. The combination of the compounds is also shown to exertgreater citrate lyase inhibiting properties than either compound alone.

Garcinol, alone, is also shown to have chemopreventive properties in ananimal tumor model.

Biological activity includes but is not limited to activity for appetitesuppression, inhibition of cytoplasmic citrate lyase, enhanced fatcatabolism, and increased anti-oxidant activity.

Hydroxycitric acid (HCA) is an alpha-hydroxy tribasic acid(1,2-dihydroxy propane -1, 2, 3-tricarboxylic acid) comprising(−)hydroxycitric acid (I), (+) hydroxycitric acid (II), (−)allo-hydroxycitric acid (III), and (+) allo-hydroxycitric acid (IV).

Garcinol is a polyisoprenylated benzophenone (PPB) derived from Garciniasp. Garcinol is a well-known anti-oxidant (i.e., emulsified garcinolsuppressed superoxide anion comparably to DL alpha-tocopherol),anti-carcinogen and also has anti-microbial properties.

Anthocyanins are one class of flavenoid compounds, which are widelydistributed plant polyphenols. There is considerable evidence thatdietary anthocyanin pigments have preventative and therapeutic roles ina number of human diseases. The chemical basis for these desirableproperties of anthocyanins is believed to be related to theirantioxidant capacity—their ability to scavenge and trap free radicalsthat damage biomolecules.

The combination of garcinol and HCA enhances bioavailability of HCA inthe cytosol and this same combination also enhances the antioxidantproperties of garcinol. This mutual enhancement of the biologicalproperties for the compounds of the invention, is referred to as “abiological effect amplification” or “Amplibio”.

Amplibio refers to a mechanism wherein either one of the compoundsserves to facilitate the biological effect of other compounds and as aresult, both biological activities are enhanced.

In many biological systems, interaction of two or more components mayresult in no interaction, antagonism, synergism or a distinct form ofsynergism called amplibio. Not all compounds that exert synergism exertamplibio action. For example, there is an advantage in combining alphalipoic acid with vitamins E and/or C, because lipoic acid recycles bothvitamins, making them more bioavailable to the body. However, bothvitamins do not enhance or otherwise improve the action of the lipoicacid. On the other hand, the results obtained by combiningpolyunsaturated fatty acids with fat soluble vitamins exemplifies anAmplibio effect: Vitamin E preserves fatty acids by preventingoxidation, while fatty acids enhance absorption of vitamin E. Anotherexample of an Amplibio effect is observed when supplemental nutrientsare combined with compounds endogenously produced by the living body:curcuminoids are known to enhance glutathione levels in the body, whichoccurs through regulating the rate of glutathione use inglutathione-free radical reaction. Elevated glutathione, in turn,increases the availability of curcuminoids in the body.

The Amplibio action of the present invention may involve the followingmechanisms:

-   -   the anti-oxidant action of garcinol/anthocyanin (enhanced by        HCA) neutralizing negative charges surrounding the cell to        facilitate transport and uptake of HCA into the cell;    -   garcinol/anthocyanin affecting fluidity of cell membranes to        facilitate transport and uptake of HCA into the cell;    -   garcinol/anthocyanin synergizing with HCA to inhibit citrate        lyase, thus lowering the threshold for such inhibition with the        HCA.

Therefore, enhancement of HCA activity is shown to be effected by itsinteraction with garcinol/anthocyanin, likewise, HCA is observed toenhance the biological potential of garcinol and anthocyanin. Thequalitative and quantitative interaction among the three compounds ofthe invention have not been described previously.

Methods for extracting and purifying HCA, garcinol and anthocyanincompounds are well known in the art.

The present invention also includes a method of preparing a compositioncomprising PPB, anthocyanin and HCA extracted from fruits of Garciniasp., the method comprising:

-   -   (a) extracting the fresh fruits of Garcinia with 8 to 1 5 parts        of C1 to C6 alcohol (e.g. methanol, ethanol, isopropyl alcohol)        by refluxing and circulating in a closed reactor until        extraction is completed;    -   (b) filtering the extracted material from the spent material;    -   (c) concentrating the extracted material to ¼–⅙ its volume under        vacuum at 540 to 600 mm/Hg;    -   (d) chilling the concentrate for approx. 20 to 30 hours at −5 to        5° C. and filtering the precipitated material in a Nutsche        filter;    -   (e) obtaining a solid residue (yield 2.5 to 4%) comprising crude        Garcinol (Assay 16–20%).    -   (f) dissolving crude Garcinol extract in a hydrocarbon solvent        such as petroleum ether, hexane or toluene (3 to 8 parts), and        extracting/partitioning with a high polar, non-miscible solvent        such as acetonitrile (2 to 4 parts, 3 to 5 times);    -   (g) separating and concentrating the polar solvent layer to ¼–⅙        its volume, and chilling at −5 to 8° C. for 12 to 16 hours;    -   (h) filtering and washing the residue with the polar solvent to        obtain yellow Garcinol (yield=0.8 to 1.5%, Assay 55 to 70%);    -   (i) adding to the extracted material of step c), molar        equivalent quantities of alkali (sodium hydroxide or potassium        hydroxide) calculated on the basis of the content of HCA (about        20 to 25%);    -   (j) heating the mixture under stirring at 80 to 95° C. for 2.5        to 5 hours to obtain yellow PPB (Garcinol);    -   (k) filtering the precipitated material (HCA salt of sodium or        potassium) using a Nutsche filter and washing with cold alcohol;    -   (l) dissolving the precipitated material in 3 to 5 parts of        water;    -   (m) adding to this aqueous solution, molar equivalent quantity        of calcium chloride and heating with stirring at 80 to 95° C.        for 4 to 6 hours;    -   (n) filtering and drying the precipitated material at 50 to        75° C. under vacuum to obtain white HCA calcium (yield=25 to        35%);    -   (o) blending the yellow PPB (Garcinol) powder obtained in        step (j) and the HCA calcium obtained in step (n) at a ratio of        1:10 to yield a composition containing HCA calcium at a minimum        of 55% and PPB (Garcinol) at a minimum of 5%; and    -   (p) optionally blending HCA calcium or any other salt of HCA,        with PPB and proantho-cyanidins in effective proportions at a        minimum of 0.5% of the proanthocyanidins.

The PPB composition of the invention can also be obtained as follows:

-   -   (a) extracting Garcinia spent fruit from suspension with toluene        and 5% methanol to obtain a paste having 20% PPB's        (Garcinol—15%; Cambogin—5%);    -   (b) upgrading the paste to obtain a composition comprising 50%        Garcinol and approx. 10–15% Cambogin which comes in the mother        liquor, and 50% Garcinol is upgraded further by one more        crystallization to yield 90% Garcinol plus 35% cambogin;    -   (c) heating a 90% Garcinol composition to 100° C. for conversion        to Cambogin of about 80% proof, the reminder being Garcinol;    -   (d) mixing the product of step (c) in equal proportion with part        of a 90% Garcinol composition to obtain a 85%–90% PPB        composition with 42–45% proof;    -   (e) obtaining a final composition with a 90% PPB in almost equal        proportion to Cambogin and Garcinol, the remainder being        cyclised products called poly isoprenylated xanthones.

The term “bioavailability” is defined as any means to enhance theeffective mechanism of a weight loss compound. By analogy, thebioavailability of calcium in the body is ultimately measured byincreased bone mass, which means that any mechanism serving the purposeof incorporating the supplemental calcium in the bone will increasebioavailability of calcium.

The term “effective amount” means a sufficient amount of compound, e.g.corresponding to HCA, garcinol or anthocyanin delivered to produce anadequate level of cytosolic citrate lyase.

A primary object of the invention is to provide a method of therapywhere the bioavailability of HCA is dramatically increased to obtaintherapeutically effective amounts of HCA in the cytosol, therebytreating the subject in need of reducing or inhibiting citrate lyase.This enhanced activity for HCA is achieved by co-administration withcompounds including but not limited to garcinol and anthocyanin.

Another object of the invention is to provide a method of weight-losstherapy where the effects of forskolin (U.S. Pat. No. 5,804,596) aredramatically increased by the presence of garcinol and, optionally,anthocyanin. The amount of forskolin is the same as taught in the '596patent, and the amounts of garcinol and anthocyanin are the same astaught herein for use with HCA.

More specifically, a weight loss and appetite suppressing effectiveamount of HCA and garcinol is 500 mg tid and 25 mg respectively, morepreferably the dose of garcinol can range from 0.01% to 10% of the HCAor other weight loss compounds like diterpene forskolin (U.S. Pat. No.5,804,596). A weight loss effective amount of HCA and anthocyanin is 500mg tid and 10 mg respectively, more preferably the dose of anthocyanincan range from 0.01% to 10% of the HCA or other weight loss compoundslike diterpene forskolin (U.S. Pat. No. 5,804,596). A weight loss andappetite suppressing effective amount of HCA, garcinol and anthocyaninis 500 mg, 25 mg and 10 mg respectively, more preferably the dose ofgarcinol and anthocyanin can range from 0.01% to 10% of the HCA or otherweight loss compounds like diterpene forskolin (U.S. Pat. No.5,804,596).

A citrate lyase inhibiting effective amount of HCA and garcinol is 500mg tid and 25 mg respectively, more preferably the dose of garcinol canrange from 0.01% to 10% of the HCA or other weight loss compounds likediterpene forskolin (U.S. Pat. No. 5,804,596). A citrate lyaseinhibiting effective amount of HCA, and anthocyanin is 500 mg tid and 10mg respectively, more preferably the dose of anthocyanin can range from0.01% to 10% of the HCA or other weight loss compounds like diterpeneforskolin (U.S. Pat. No. 5,804,596). A citrate lyase inhibitingeffective amount of HCA, garcinol and anthocyanin 500 mg, 25 mg and 10mg respectively, more preferably the dose of garcinol and anthocyanincan range from 0.01% to 10% of the HCA or other weight loss compoundslike diterpene forskolin (U.S. Pat. No. 5,804,596).

A fat catabolizing and lean body mass enhancing effective amount of HCAand garcinol is 500 mg tid and 25 mg respectively, more preferably thedose of garcinol can range from 0.01% to 10% of the HCA or other weightloss compounds like diterpene forskolin (U.S. Pat. No. 5,804,596). A fatcatabolizing and lean body mass enhancing effective amount of HCA andgarcinol is 500 mg tid and 25 mg respectively, more preferably the doseof garcinol can range from 0.01% to 10% of the HCA or other weight losscompounds like diterpene forskolin (U.S. Pat. No. 5,804,596). A fatcatabolizing and lean body mass effective amount of HCA, garcinol andanthocyanin is 500 mg, 25 mg and 10 mg respectively, more preferably thedose of garcinol and anthocyanin can range from 0.01% to 10% of the HCAor other weight loss compounds like diterpene forskolin (U.S. Pat. No.5,804,596).

The therapeutic weight-loss effects of HCA and garcinol, in combination,and the role of garcinol as chemopreventive agent are described by wayof the following non-limiting examples.

EXAMPLE 1

Administration of garcinol and HCA to two strains of mice, SKH-1 andCF-1 respectively, resulted in significantly less total body weight andabdominal fat gain, as compared to control, chow-receiving animals andthe groups of animals receiving either garcinol or HCA alone. Inaddition, dietary administration of garcinol caused significantreduction in aberrant colonic crypt formation (AC is considered apre-malignant condition) in CF-1 mice as compared to the animals fedcontrol diet or diet containing HCA or garcinol alone. Interestingly,the weight-gain preventive effect of garcinol and HCA combination wasaccomplished despite that the garcinol plus HCA animals had higher foodand water consumption than the control, garcinol and HCA groups.

TABLE 1 Effect of oral administration of garcinol (GAR), hydroxycitricacid (HCA) and combination of garcinol and hydroxycitric acid on bodyweight, food and fluid consumption in SKH-1 mice Group 5 weeks 7 weeks10 weeks Body weight (gm; Mean ± SE) 1. Control 32.3 ± 0.67 34.5 ± 0.9937.0 ± 1.49 2. 0.05% GAR 32.8 ± 0.37 33.9 ± 0.14 36.4 ± 0.55 3. 1% HCA32.5 ± 0.42 34.5 ± 0.59 36.8 ± 0.33 4. (2) + (3) 31.2 ± 0.20 33.5 ± 0.5834.9 ± 0.88 Food consumption (gm/mouse/day; Mean ± SE) 1. Control 5.25 ±0.33 5.09 ± 0.28 5.16 ± 0.25 2. 0.05% GAR 4.98 ± 0.17 5.09 ± 0.46 5.26 ±0.17 3. 1% HCA 5.74 ± 0.13 6.49 ± 0.07 6.93 ± 0.21 4. (2) + (3) 6.55 ±0.20 8.03 ± 1.45 9.31 ± 1.12 Water consumption (ml/mouse/day; Mean ±SE) 1. Control 3.75 ± 0.04 3.42 ± 0.34 3.80 ± 0.22 2. 0.05% GAR 3.66 ±0.07 3.63 ± 0.05 3.73 ± 0.17 3. 1% HCA 3.72 ± 0.08 3.74.5 ± 0.04   3.84± 0.09 4. (2) + (3) 3.68 ± 0.08 3.86 ± 0.08 3.94 ± 0.08

TABLE 2 Effect of 10 week oral administration of garcinol (GAR) andhydroxy- citric acid (HCA) on azoxymethane (AOM)-induced formation ofaberrant colonic crypts (AC) and accumulation of fat in abdomen in CF-1mice Body weight AC parametrial retroperiton- Group (gm) per colon fat(gm) eal fat (gm) 1. Control 38.8 ± 1.52 11.4 1.33 ± 0.19 0.95 ± 0.11 2.0.05% GAR 37.3 ± 1.07  7.8 (31.6%) 1.21 ± 0.18 0.85 ± 0.01 3. 1% HCA37.9 ± 0.66  7.9 (30.7%) 1.24 ± 0.06 0.79 ± 0.04 4. (2) + (3) 36.1 ±0.48  8.5 (25.4%) 0.95 ± 0.10 0.70 ± 0.13These data suggest a broad biological mechanism for garcinol as anadjuvant to HCA involving regulation of body weight, body compositionand body metabolic rate.

EXAMPLE 2

The role of garcinol as a chemopreventive agent has been confirmed in anexperiment in which CF-1 mice were treated topically with theinflammation and tumor promoting agent, TPA.

TABLE 3 Effects of garcinol (GAR) on TPA-induced ear edema* NumberWeight per Percent Inhibition Treatment of mice punch (gm) (%) 1. TPA (1nmol) 5 15.5 ± 0.82 — 2. TPA + GAR (0.1 umol) 4 12.4 ± 0.81 38.5 3.TPA + GAR (0.5 umol) 4 9.25 ± 0.66 77.5 *Female CF-1 mice were treatedtopically with TPA applied to ear's skin with or without garcinol. Fivehours later, the mice were killed and ear punches were weighted. Dataare expressed as the mean ± SE.

These data suggest a biological mechanism for garcinol as achemopreventative agent.

EXAMPLE 3

Clinical Studies on Weight-Loss Potential of Garcinol and HCA

The combination consisting of 500 mg of calcium salt of HCA and 25 mg ofgarcinol (NC) was evaluated in a double-blind, 12 week clinical studyagainst the formula containing 500 mg of calcium salt HCA (C). The studywas performed on 46 overweight female volunteers (BMI greater than 25).Participants were instructed to take one capsule of eitheractive-containing or placebo formula three times a day, half an hourbefore a meal. Each participant was asked to maintain her previous dailyphysical exercise and eating habits. In addition, physical activity wasmonitored based on a questionnaire before and during the trial. Theparticipants were evaluated at the baseline, weeks 2, 4, 6, 8, 10 and12. The following clinical parameters were evaluated at each visit:total body weight, body composition by the bioelectric impedance method,self-assessed appetite and energy levels, pulse rate and blood pressure.During the 12 week trial, the mean values in group NC for body weightand fat content significantly decreased, whereas lean body mass andtotal body water significantly increased compared to the baseline valuesand C group values. The appetite levels were significantly less in theNC group than the C group, whereas energy levels were equally increasedin both study groups as compared to the baseline. No subjective orobjective adverse effects were reported in the course of this study. Thepulse rate, systolic and diastolic blood pressure were maintained at thesame level throughout the study.

NC is statistically more effective than C in reducing total body weightand body mass index; reducing body fat; increasing lean body mass andcontent of body water; and reducing levels of appetite perception. Theenergy levels of subjects were not enhanced by regimens in both groups,and NC and C did not produce subjective or objective side effects.

INDUSTRIAL APPLICATIONS

The combined data from animal and clinical studies indicate that theinvention is more effective than HCA alone weight-loss therapy, and thatin addition to fat loss, the inventive composition invention alsoimproves lean body mass leading to improved body composition.Maintaining or improving lean body mass while shedding extra pounds ofadipose tissue has been advocated by the Centers for Disease Control.Lean body mass is important because it has been recently recognized asan independent, positive predictor of one's cardiovascular health andoverall health. The chemopreventive action of garcinol in the animalexperimental model may also be intricately related to the role thecompound has in improving body composition.

REFERENCES

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1. A method for increasing fat catabolism in a subject in need of sucheffect comprising administering to said subject a fat catabolizingeffective amount wherein the active ingredients in said fat catabolizingamount consists essentially of hydroxycitric acid and garcinol ofhydroxycitric acid and garcinol.
 2. The method of claim 1, wherein thefat catabolizing effective amount of hydroxycitric acid and garcinolfurther comprises anthocyanin.
 3. The method of claim 1, wherein thehydroxycitric acid is extracted from fruits of Garcinia sp. with organicsolvents.
 4. The method of claim 1, wherein the garcinol is extractedfrom fruits of Garcinia sp. with organic solvents.
 5. The method ofclaim 2, wherein the hydroxycitric acid (HCA), garcinol and anthocyaninare extracted from fruits of Garcinia sp. with organic solvents.
 6. Themethod of claim 1, wherein the hydroxycitric acid is extracted fromfruits of Garcinia sp. by CO₂ supereritical extraction.
 7. The method ofclaim 1, wherein the garcinol is extracted from fruits of Garcinia sp.by CO₂ supercritical extraction.
 8. The method of claim 2, wherein thehydroxycitric acid, garcinol and anthocyanin are extracted from fruitsof Garcinia sp. by CO₂ supercritical extraction.
 9. The method of claim1, wherein the fat catabolizing effective amount of hydroxycitric acidand garcinol also increases lean body mass.
 10. The method of claim 2,wherein the fat catabolizing effective amount of hydroxycitric acid,garcinol and anthocyanin also increases lean body mass.
 11. The methodof claim 1, wherein the fat catabolizing effective amount ofhydroxycitric acid and garcinol also promotes weight loss.
 12. Themethod of claim 2, wherein the fat catabolizing effective amount ofhydroxycitric acid, garcinol and anthocyanin also promotes weight loss.13. The method of claim 1, wherein the fat catabolizing effective amountof hydroxycitric acid is 500 mg.
 14. The method of claim 1, wherein thefat catabolizing effective amount of garcinol is 25 mg.
 15. The methodof claim 2, wherein the fat catabolizing effective amounts ofhydroxycitric acid, garcinol and anthocyanin are 500 mg, 25 mg, and 10mg respectively.
 16. The method of claim 1, wherein the percentage ofgarcinol in the fat catabolizing effective amount of hydroxycitric acidand garcinol ranges from 0.01% to 10% relative to hydroxycitric acid.17. The method of claim 2, wherein the percentage of garcinol in the fatcatabolizing effective amount of hydroxycitric acid, garcinol andanthocyanin ranges from 0.01% to 10% relative to hydroxycitric acid.